**3. Materials and methods**

Previously investigated targets, the protein kinase domain of human Ephrin Receptor Tyrosine Kinase A3 (EphA3) (Davis *et al.*, 2008) and the *Pseudomonas aeruginosa*  alkylhydroperoxidase D protein (PA0269) (McGrath *et al.,* 2007) were selected as model proteins to demonstrate the feasibility of crystal growth and *in situ* data collection using the X-CHIP. Following several rounds of on-chip optimization, both projects were crystallized at conditions similar to those in the cited literature. EphA3, at 15 mg ml-1, and native PA0268, at 10 mg ml-1, were crystallized using a 1:1 (vol/vol) with 0.2M (NH4)2SO4, 0.1M Hepes pH 7.5, 25% PEG3350 and 0.8M (NH4)2SO4, 0.1M sodium citrate pH 4.0 respectively. For crystallization drop set up, a protein sample volume of 200-250nl was mixed with an equal volume of precipitant solution and then covered by an mineral oil volume of 0.75-1.25μl; all solutions were dispensed with a Gilson P2 pipette. Prior to set up, the chip was inserted into the holder, which was afterward covered with a lid to prevent contamination. Chips were stored for 1-4 weeks at room temperature without significant evaporation and transported by road to the synchrotron beamline. Crystals both from stored chips as well as from chips set up at the synchrotron were used to collect diffraction data sets.

In-house data sets were collected on a *Rigaku FR-E* Superbright rotating anode X-ray source equipped with a *Rigaku R-AXIS HTC* image plate as a detector (Rigaku, The Woodlands, TX, USA). The synchrotron data sets were collected on the *IMCA-CAT ID-17* beamline at the Advanced Photon Source (APS) facility with appropriate beam attenuation, using the *Pilatus 6M* detector (Dectris Ltd., Baden, Switzerland). The X-CHIP was manually mounted onto the goniometers as shown in Fig. 2*d*. Individual samples were initially optically centered, and then centered using diffraction to refine crystal position. In both cases the cryo-stream was blocked and data collection was performed at room temperature (~298K).
